三维子宫组织的构建及形态学观察

The Construction and Morphology Observation of Three-dimensional Uterine Tissue

本研究旨在体外构建含有3种细胞的三维子宫组织。体外成功分离和培养兔子宫内膜细胞和平滑肌细胞,将3代以内的平滑肌细胞以1×107/mL与3 mg/mLⅠ型鼠尾胶原和Matrigel(4∶1)的混合物混合,迅速接种在自制的模具中,置于37℃、体积分数5%CO2培养箱中待其凝固,以同样的方法接种1×107/mL子宫内膜基质细胞,待其凝固后接种上皮细胞悬液,将其置于37℃,体积分数5%CO2培养箱中培养。采用HE染色和免疫组织化学染色方法观察和鉴定三维子宫组织。结果表明,在体外成功地构建了含有3种细胞结构的类似于在体子宫组织的三维子宫组织;HE染色观察,在静态外力作用下,培养10 d的子宫组织细胞呈明显的按拉伸方向排列;细胞在细胞外基质中自由移动,上皮层呈弯曲状;免疫组织化学染色可以观察到具有3层细胞结构的子宫组织,弯曲的上皮层细胞呈柱状。可见子宫内膜细胞和平滑肌细胞能在细胞外基质中生长;建立了一套由3种细胞组成的类似于在体子宫组织结构的三维子宫培养体系;为胚胎植入机理的研究提供良好的体外模型。

Three-dimensional uterine tissue was reconstructed in vitro, which was composed of 3 kinds of cells. Rabbit endometrial cells and smooth muscle cells were successfully isolated and cultured in vitro. Smooth muscle cells less than 3 generations （ 1 × 10^7 cells/mL） were mixed with the mixture of 3 mg/mL collagen I solution derived from mouse tail tendon and Matrigel gel（4 ： 1）, and were inoculated quickly into self-made moulds, the cell-gel mixture solidified for 30 min in 5% CO2 at 37℃. Endometrial stromal cells （ 1×10^7 cells/mL） were seeded in the mixture by the same ways, the suspension of endometrial epithelial cells was inoculated on the gel when it was solidified; The tissue were cultured in 5% CO2 at 37℃. HE staining and immunohistoehemieal staining were used to analyze and identify the three-dimensional uterine tissue. The resuits indicated that three-dimensional uterine tissue was successfully reconstructed in vitro, which Was composed of three kinds of cells and whose structure was similar to native uterine tissue. HE staining analyses showed that reconstructed uterine tissue was stretched along the stretching direction on 10 d, cells moved freely in extracellular matrix; The epithelial layer showed curving. Immunohistochemical staining revealed that reconstructed uterine tissue distributed three layers of cells; Cells of coiled epithelial layer showed column aspect. Obviously, endometrial cells and smooth muscle cells successfully grow in extracellular matrix; The cultured sys-tern of three dimensional uterine was reconstructed in vitro, composed of three kinds of cells and similar to native uterine sible for research on the mechanism of embryo implantation three-dimensional uterine tissue tissue. The established model is was feasible for research on the mechanism of embryo implantation.